foods
Article
Identification of Umami Taste in Sous-Vide Beef by
Chemical Analyses, Equivalent Umami
Concentration, and Electronic Tongue System
Young-Hwa Hwang 1,† , Ishamri Ismail 2,3,† and Seon-Tea Joo 1,2, *
1
2
3
*
†
Institute of Agriculture & Life Science, Gyeongsang National University, Jinju 52828, Korea;
philoria@gnu.ac.kr
Division of Applied Life Science (BK21+), Gyeongsang National University, Jinju 52828, Korea;
ishamriismail@unisza.edu.my
Faculty of Bioresources and Food Industry, Universiti Sultan Zainal Abidin, Besut Campus,
Terengganu 22200, Malaysia
Correspondence: stjoo@gnu.ac.kr; Tel.: +82-55-772-1943
These authors contributed equally to this work.
Received: 16 January 2020; Accepted: 24 February 2020; Published: 26 February 2020
Abstract: Behaviour of umami compounds that are associated with non-volatile compounds on slow
cooking regimes remains less explored. This study aims to assess the ability of the electronic tongue
system on the umami taste from sous-vide beef semitendinosus. The identification was based on the
taste-enhancing synergism between umami compounds 5’-nucleotides (IMP, GMP, AMP, inosine, and
hypoxanthine) and free amino acids (glutamic and aspartic acid) using the estimation of equivalent
umami concentration (EUC) and electronic tongue system. Sous-vide cooked at 60 and 70 ◦ C for
6 and 12 h and cooked using the conventional method at 70 ◦ C for 30 min (as control) were compared.
The temperature had a significant effect on 5’-nucleotides, but aspartic and glutamic acid were not
influenced by any treatments applied. Sous-vide cooked at 60 ◦ C tended to have higher inosine and
hypoxanthine. Meanwhile, desirable 5’-nucleotides IMP, AMP, and GMP were more intensified at the
temperature of 70 ◦ C. The principal component analysis predicted a good correlation between EUC
and the electronic tongue, with sous-vide at 70 ◦ C for 12 h presenting the most umami. Therefore,
the electronic tongue system is a useful tool in food processing, particularly in determining complex
sensory properties such as umami, which cannot be evaluated objectively.
Keywords: umami; sensory; sous-vide; electronic tongue; IMP; 5’-nucleotides
1. Introduction
Moist heat cooking is the standard cooking method for tough cut meats such as semitendinosus,
semimembranosus, and biceps femoris [1]. Sous-vide is one of the examples of moist heat cooking.
This cooking method often uses lower cooking temperatures (<95 ◦ C) and longer cooking times as
a function to minimize surface drying and to bring advantages to a gastronomic outcome [2,3]. The
primary benefit of using sous-vide cooking is that the end-product can be precisely and robustly
controlled by selecting suitable cooking conditions (i.e., temperature and time).
From our previous study on tough cuts of beef semitendinosus [4], we noticed prolonged cooking
at 60 and 70 ◦ C produces lower shear stress, with sous-vide cooked at 60 ◦ C presenting better water
retention and color properties. After this finding, curiosity about the palatability of meat led to the
exploration of a taste study. Roldán et al. [5] observed that flavor from sous-vide lamb was affected
by the temperature and time of cooking. They found that lamb cooked at low or moderate cooking
conditions (60 ◦ C for 6 and 24 h or at 80 ◦ C for 6 h) is more likely to have lipid-derived flavor compounds.
Foods 2020, 9, 251; doi:10.3390/foods9030251
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Meanwhile, at moderately high temperature for long times (80 ◦ C for 24 h) promoted a desirable
meaty flavor due to the formation of volatile compounds from Strecker degradation. Besides, previous
literature has also evidenced that sous-vide cooking preserves sensory attributes, which minimize
the loss of volatile aromatic compounds, resulting in the palatability of cooked meat [5–7]. However,
information regarding umami taste and non-volatile compounds is still limited.
Umami is an essential sensory feature of meat and plays a significant role in palatability.
The umami taste is unique with savory, brothy, or beefy characterizing this broad taste. Several
numbers of substances impart the umami sensation, predominantly amino acid glutamate and
5’-nucleotides (mainly inosine-5’-monophosphate (IMP) and guanosine-5’-monophosphate (GMP)) [8].
Also, adenosine-5’-monophosphate (AMP) and aspartic acid, to a lesser extent, contribute to the umami
attribute [9]. However, the taste of umami is not clear as compared to sweet and salty because it
is formed from the chemical reaction between substances. Although umami is the typical taste of
monosodium glutamate (MSG) [10], the MSG taste itself is not very palatable [11]. Yamaguchi and
Ninomiya [11] found that the apparent umami taste appears when IMP and glutamate are present in
the food system, likely due to the synergistic between both substances.
The mechanical instrument to measure texture properties are common in meat quality but unusual
for sensory measurement. Currently, the sensory features also can be measured using an automated
sensing machine, namely an electronic tongue. The taste sensor can measure specific tastes objectively
and uniformly [12]. Electronic tongue ability is not only limited to evaluate saltiness, sourness,
bitterness, and sweetness but also complex tastes such as umami and astringent. Moreover, this taste
sensor is highly correlated to human sensory scores, and it even offers satisfying taste results [13].
Therefore, the objective of this study is to examine the ability of an electronic tongue on the umami
attributes at different cooking temperatures (60 and 70 ◦ C) and times (6 and 12 h) of beef semitendinosus
as a comparison to the conventional cooking method at 70 ◦ C for 30 min. The factors that contribute to
umami, such as 5’-nucleotides and free amino acids, and their synergism effect are also investigated.
2. Materials and Methods
2.1. Raw Materials and Sampling
Ten semitendinosus muscles from Hanwoo beef were purchased from a local meat processing plant
at 36 h postmortem (25–30 months old; quality grade 3). The fresh weight of the beef muscles was
1803 ± 550 g. The experiment was randomly divided into 6 and 12 h groups, with each group allocated
five muscles (n = 5). Before heat treatment, the beef was cut into 2.5 cm thick slices (weight 195–211 g),
with the distal and proximal ends discarded. Only two slices were taken randomly from each muscle
and assigned as treatments 60 and 70 ◦ C. This procedure was applied to both groups of 6 and 12 h.
Then, the muscle was placed in an individual vacuum package, and they were kept refrigerated at
4 ◦ C overnight.
At 48 h postmortem, the vacuum-packed beef was cooked at 60 and 70 ◦ C for 6 and 12 h using
a Sous Vide Travellortech Precision Cooker Immersion Circulator (Travellortech, Ilion, NY, USA).
For the conventional cooking method (labeled as a control), the five slices from different muscles were
chosen randomly from groups 6 and 12 h (three slices from the 6 h group and two slices from the 12 h
group; n = 5). Subsequently, the muscles were packed without vacuum inside a polypropylene plastic
packaging and cooked at 70 ◦ C for 30 min using a water bath (JSWB-22T, 30 L, Gongju, South Korea).
Since the meat in a conventional cooking method such as braising is subjected directly to the boiling
water, it often produces imbalance outcomes such as flavorless dry meat and flavorful released juice.
Thus, in the current study, we modified the method of conventional braising (control treatment) by
cooking the beef in a package at a well-done temperature (70 ◦ C) and limited the cooking time (30 min)
as a function to avoid vigorous effects on the meat. This minimizes the loss of flavor inside a bag
and preserves the taste compounds of meat. The temperature of water in the bath for sous-vide and
conventional cooking was monitored using thermocouples (Testo, model 735–2, Lenzkirch, Germany).
Foods 2020, 9, 251
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Immediately after cooking, the packed-meat was transferred into icy cold water for one hour and then
kept under refrigeration at 4 ◦ C overnight before analysis.
2.2. 5’-Nucleotides Measurement
The measurement of 5’-nucleotides were carried out by high-performance liquid chromatography
(HPLC) based on method of Jayasena et al. [14]. Then, 5’-nucleotides: adenosine-5’-monophosphate
(AMP), inosine-5’-monophosphate (IMP), guanosine-5’-monophosphate (GMP), inosine, and
hypoxanthine were determined. Five grams of cooked meat was minced and homogenized using a
high-speed homogenizer (IKA, model T25D, Staufen, Germany), with cold 20 ml 0.5 M perchloric acid
at 10,000 rpm for 15 s. Homogenate was centrifuged at 3000× g at 4 ◦ C for 10 min and the supernatant
was filtered using filter paper No. 1 (Whatman Ltd., Little Chalfont, UK). After filtration, the residuum
again was mixed and vortexed with 10 mL 0.5 M perchloric acid and transferred into the same tube by
filtration with filter paper No. 1. The clear filtrate was adjusted to pH 6.0 with cold 5 N potassium
hydroxide (KOH) and centrifuged at 3000× g at 4 ◦ C for 10 min to allow the salt precipitate to the
bottom of the tube. After centrifugation, the pH-adjusted supernatant was filtered with filter paper
No. 4 (Whatman Ltd., Little Chalfont, UK) and brought to a volume of 50 mL by adding cold 0.5 M
perchloric acid (pH 6.0 adjusted with 5 N KOH). The supernatant was filtered through a 0.45 µm
polytetrafluoroethylene syringe filter (SF13-HLB45-M, Futecs Co. Ltd., Daejeon, South Korea) into a
2-mL wide opening screw-top glass HPLC vial (PN: 5182-0557, Agilent, Santa Clara, CA, USA).
The filtrate was analyzed by a Hewlett-Packard HPLC system (Model 1100, Germany) using UV
detection at 260 nm. Separation of the 5’-nucleotides was achieved on a C18 column (4.6 × 250 mm,
5 µm particle size, Acclaim TM 120, Thermo Scientific, Waltham, MA, USA) with the temperature of
the column set at 25 ◦ C. Mobile phase A was 0.06 M dipotassium hydrogen phosphate and 0.04 M
potassium dihydrogen phosphate at pH 7.0. Mobile phase B was a mixture of HPLC-grade water
and methanol (20:80 v/v). The separation was performed in a gradient elution mode with a constant
flow rate at 0.8 ml/min, stop time at 16 min, and post time at 5 min. The gradient program is shown
in Table 1. The injection (10 µL) with a needle wash program was used after each injection, using a
washing mixture of water and methanol (1:1 v/v). Identification of 5’-nucleotides was carried out by
comparing the retention time in samples with their retention time in standard (AMP, IMP, GMP, inosine,
and hypoxanthine; Sigma-Aldrich Co., St. Louis, MO, USA). The concentrations were quantified based
on the calibration curve and data was expressed as g/100 g of sample.
Table 1. 5’-nucleotides gradient elution conditions.
Time (min)
Mobile Phase B (%)
0
5
7
8
9
16
2
4
4
30
0
0
2.3. Glutamic and Aspartic Acid Determination
Glutamic and aspartic acid were analyzed and identified using HPLC based on the method of
Mæhre et al. [15]. Free amino acids were prepared with 0.4 g of minced meat and hydrolyzed under
nitrogen gas at 110 ◦ C for 16 h with 15 mL of 6 M HCL. The aliquots (100 µL) was re-dissolved with
Mili-Q water (1 mL) after evaporating to dryness with nitrogen gas. Then, they were filtered through a
0.45-µm polytetrafluoroethylene syringe filter. The standard solution and hydrolyzed samples were
derivatized with O-pthaldialdehyde (OPA) (PN: 5061-3335, Agilent, Santa Clara, CA, USA) after mixing
with borate buffer at pH 10.4 using an autosampler. The autosampler injector program is shown in
Table 2. Standard/sample solution was injected through an Agilent ZORBAX Eclipse AAA column (4.6
Foods 2020, 9, 251
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× 150 mm, 5 µm particle size, Agilent, USA) at 338 nm and column compartment temperature was
set at 40 ◦ C (left and right). Mobile phase A contained 40 mm NaH2 PO4 pH 7.8 (adjusted with 10 N
NaOH), and mobile phase B contained a mixture of acetonitrile, methanol, and water (45:45:10 v/v/v).
The separation was performed using a gradient elution program, as shown in Table 3, with a flow rate
at 1.5 mL/min, stop time at 25 min, and post time at 5 min. The individual peaks of glutamic and
aspartic acid were identified based on the retention times of standard compounds.
Table 2. Injector program.
Line
Function
Amount
Reagent
Vial
Position
1
Draw
2.5 µL
Borate buffer pH 10.4
1
2
Draw
0.5 µL
Sample
11
3
Mix
3 µl in air, max speed 2×
4
Wait
0.5 min
5
Draw
0 µL
HPLC-grade water (for needle
was using water in uncapped vial)
2
6
Draw
0.5 µL
O-pthaldialdehyde (OPA)
3
7
Mix
3.5 µL in air, max speed 6×
8
Draw
0 µL
HPLC water (for needle was using
water in uncapped vial)
2
9
Mix
4 µL in air, max speed 6×
10
Draw
0 µL
Acetonitrile
5
11
Draw
32 µL
HPLC-grade water
4
12
Mix
18 mL in air, max speed 2×
13
Inject
Table 3. Free amino acids gradient elution conditions.
Time (min)
Mobile Phase B (%)
0
0.5
20
20.1
23.5
23.6
25
2
2
57
100
100
2
0
2.4. Equivalent Umami Concentration (EUC)
The estimation of umami can be measured based on the concentration of monosodium glutamate
(MSG) by EUC value (mg MSG/g), which is equivalent to the umami intensity. The synergy effect
between umami amino acids and 5’-nucleotides is represented by the following equation [16]:
Y=
X
ai bi + 12.18(
X
ai bi )(
X
aj bj )
(1)
where Y is the EUC (mg MSG/g); ai is the concentration (mg/g) of each umami amino acid (aspartic or
glutamic acid); aj is the concentration (mg/g) of each umami 5’-nucleotide (IMP, GMP, or AMP); bi is
the relative umami concentration (RUC) for each umami amino acid to MSG (glutamic acid: 1 and
aspartic acid: 0.077); bj is the RUC for each umami 5’-nucleotide to IMP (IMP: 1, GMP: 2.3, and AMP:
0.18); and 12.18 is a synergistic constant based on the concentration of mg/g used.
Foods 2020, 9, 251
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2.5. Umami Determination by Electronic Tongue
The electronic tongue system (INSENT SA402B, Tokyo, Japan) was used to determine sous-vide
taste information. The instrument included sensor arrays (reference and lipid/membrane electrodes),
an autosampler, an electronic unit for data acquisition, and a chemometric data analysis software
package. This system was composed of five taste sensors, and each sensor was fixed with a unique
2020,sensors
9, 251
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artificial lipid.Foods
The
were named as AAE (to detect umami substances), CA0 (to5 ofdetect
sourness
substances), C00
(to detect bitter substances), AE1 (to detect astringent substances), and CT0 (to detect
2.5. Umami Determination by Electronic Tongue
saltiness substances).
The inner solution consisting of 3.33 M potassium chloride in saturated silver
The electronic tongue system (INSENT SA402B, Tokyo, Japan) was used to determine sous-vide
chloride solution
was
provided
by INSENT
Inc.
(Atsugi-shi,
Japan)
and used forelectrodes),
the sensor arrays of
taste information. The instrument
included
sensor
arrays (reference
and lipid/membrane
an
autosampler,
an
electronic
unit
for
data
acquisition,
and
a
chemometric
data
analysis
software solution as
lipid/membrane and reference electrode [13]. All sensor was preconditioned in a standard
package. This system was composed of five taste sensors, and each sensor was fixed with a unique
described byartificial
Woertz
et al. [17], and a sensor check was done routinely before every measurement to
lipid. The sensors were named as AAE (to detect umami substances), CA0 (to detect sourness
ensure the system
works
substances),
C00 properly.
(to detect bitter substances), AE1 (to detect astringent substances), and CT0 (to detect
saltiness substances).
The inner solutionextraction
consisting of was
3.33 Mbased
potassium
in saturated
silver
The preparation
of the water-soluble
on chloride
the method
of Hwang
et al. [18].
chloride solution was provided by INSENT Inc. (Atsugi-shi, Japan) and used◦ for the sensor arrays of
Minced meatlipid/membrane
(50 g) was and
stirred
in
200
mL
hot
distilled
water
at
95
C
(1:4
w/v)
for
10
min
before
reference electrode [13]. All sensor was preconditioned in a standard solution
centrifugation
at 3000×
for 10
min.
filtered
through
filter paper No. 1
as described
bygWoertz
et al.
[17], The
and a supernatant
sensor check waswas
done then
routinely
before every
measurement
to ensure
theChalfont,
system works
properly.
(Whatman Ltd.,
Little
UK)
prior to analysis by the electronic tongue.
The preparation of the water-soluble extraction was based on the method of Hwang et al. [18].
The measurement
procedure was as follows: first, the sensor arrays were immersed in the
Minced meat (50 g) was stirred in 200 mL hot distilled water at 95°C (1:4 w/v) for 10 min before
reference solution
(innerat solution)
obtain
the membrane
Vr . filter
Second,
arrays
centrifugation
3000·g for 10tomin.
The supernatant
was thenpotential,
filtered through
paper the
No. sensor
1
(Whatman
Little Chalfont,
UK)
to analysis
by the electronic
tongue. V . Third, the sensor arrays
were immersed
in theLtd.,
sample
solution
toprior
obtain
the membrane
potential,
s
The measurement procedure was as follows: first, the sensor arrays were immersed in the
were rinsed lightly
with
the
reference
solution
and,
subsequently,
they
were
immersed again in the
reference solution (inner solution) to obtain the membrane potential, Vr. Second, the sensor arrays
reference solution
to obtain
the
potential,
Vobtain
the sensor
arrays
were
with an alcohol
were immersed
in the
sample
solution to
the membrane
potential,
Vs. Third,
the rinsed
sensor arrays
r ’. Finally,
with
the reference
solution and,
subsequently,
they were
immersed again
in the
solution (100 were
mmrinsed
KCl, lightly
10 mm
KOH,
30% ethanol)
to remove
adsorbed
substances
before
measuring the
reference solution to obtain the potential, Vr’. Finally, the sensor arrays were rinsed with an alcohol
next sample. solution
The relative
CPA 30%
value
(change
of membrane
potential
by adsorption)
(100 mmvalue
KCl, 10and
mm KOH,
ethanol)
to remove
adsorbed substances
beforecaused
measuring
are shown in the
Figure
1. The The
sequential
measurement
was(change
repeated
four times
for each
sample,
with the
next sample.
relative value
and CPA value
of membrane
potential
caused
by
adsorption)
are
shown
in
Figure
1.
The
sequential
measurement
was
repeated
four
times
for
each
first measurement was discarded to enable conditioning of the sensors. Only the umami response
sample, with the first measurement was discarded to enable conditioning of the sensors. Only the
output was taken
presented
intaken
this and
paper.
umamiand
response
output was
presented in this paper.
Figure 1. Measurement diagram. The relative value is the initial taste at sensory evaluation, including
Figure 1. Measurement diagram. The relative value is the initial taste at sensory evaluation, including
sourness, saltiness, and umami. The CPA value is the adsorption of bitter and astringent substances.
sourness, saltiness,
umami.
The
CPAbevalue
is the
of bitter
and
astringent
Note: theand
reference
solution
should
tasteless
or atadsorption
low concentration
than the
tested
sample. substances.
Adoptedsolution
from Kobayashi,
Habara,
Ikezazki, or
Chen,
Naito,concentration
and Toko [13]. than the tested sample. Adopted
Note: the reference
should
be tasteless
at low
from Kobayashi, Habara, Ikezazki, Chen, Naito, and Toko [13].
2.6. Data Analysis
The results of umami related compounds and umami taste evaluation were recorded as mean
and standard error of mean (SEM). All assays were performed in triplicate. The interaction between
temperatures and times (2 temperatures × 2 times) were performed by analysis of variance general
Foods 2020, 9, 251
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linear model (GLM) using SPSS version 23 (IBM Corp., SPSS Inc., Chicago, IL, USA). The multiple
comparisons between control and sous-vide treatments were done at the 5% level by Tukey’s test.
Principal component analysis (PCA) was used to test the discrimination performances between cooked
samples and tested variables. The first two components (F1/F2) were used to develop the variable
and observation plot, and the correlation was observed from the two-dimensional map. PCA was run
using XLSTAT software (Addinsoft INC, Brooklyn, NY, USA).
3. Results and Discussion
3.1. 5’-Nucleotides
The concentration of five 5’-nucleotides were analyzed in the sous-vide cooked meat. As presented
in Table 4, all 5’-nucleotides were significantly affected by temperature, while the effect of time was only
not significant on the IMP and hypoxanthine (p = 0.061 and 0.968, respectively). IMP was comprised of
higher composition than the other 5’-nucleotides and it was expected to contribute mainly to the umami
threshold [16,19,20]. Besides IMP, other 5’-nucleotides such as GMP and AMP were also important
components in imparting umami taste [21]. In all treatments, beef sous-vide cooked for 12 h at 70 ◦ C
showed significantly higher IMP, GMP, and AMP, with a GMP value double of that of 6 h of cooking at
the same temperature. Unexpectedly, sous-vide cooked at 60 ◦ C, regardless of cooking time, recorded
a lower concentration of IMP, GMP, and AMP, but instead, it led to significantly higher contents of
inosine and hypoxanthine. The data presented here are similar to those reported for cooked meat
samples in previous literature for pork [9], goat [22], and beef [23], which found lower IMP and higher
inosine at 60 ◦ C, although there are some differences.
As shown in Table 4, the content of IMP of control was decreased by at least 6% after 30 min of
cooking (initial IMP = 49.63 g/100 g), while the content of IMP for sous-vides decreased by 20%–28%
and 38–39% after prolonged cooking at 70 and 60 ◦ C, respectively. The degradation of IMP evidenced
the significant effect of temperature was more prominent and can be elaborated further by either two
mechanisms: (i) dephosphorylation of IMP into inosine; or (ii) IMP and ribose as flavor precursors
involve in a number of secondary reactions to yield heterocyclic volatiles compounds [24,25]. This could
be explained by which reaction takes place depending on the temperature applied [24]. The reason
why 60 ◦ C cooking has lower IMP values and 70 ◦ C cooking has higher IMP values could be elaborated
by enzymatic activity, as suggested by Tomioka et al. [26]. In our previous experiment, we investigated
the proteolytic activity between temperatures and times of sous-vide cooked beef semitendinosus, and
we found that at 60 ◦ C cooking had higher proteolytic activity than at 70 ◦ C. This enzyme activity did
not change even after 12 h of cooking period [4]. Similarly, Ishiwatari, Fukuoka, Hamada-Sato, and
Sakai [23] studied the kinetic model of cooked meat and they found that cooked meat heated above
than 60 ◦ C could slow down the activity of the enzyme. They also found that complete deactivation of
the IMP-decomposition enzyme activity was reached at 64.1 ◦ C.
3.2. Aspartic Acid and Glutamic Acid
Among all the amino acids, aspartic and glutamic acid play a primary role in contributing to
umami taste [27]. Therefore, in this paper, we focused only on these amino acids rather than profiling
of all amino acids in the sous-vide cooked meat. Interestingly, both aspartic and glutamic acids did not
have statistically significant differences at any temperature and time of sous-vide treatment. When
viewed at initial values (aspartic acid = 0.49 g/100 g and glutamic acid = 0.59 g/100 g), all the sous-vide
treatments contributed to an increase in amino acids of aspartic and glutamic. The present data were
in tandem to the finding by Rotola-Pukkila, Pihlajaviita, Kaimainen, and Hopia [9], who also found
no difference in both amino acids at different temperatures and times on pork loins. According to
Yamaguchi and Ninomiya [11], the detection of glutamate was noticeably lowered in the presence of
IMP, likely due to the synergistic effect. However, the results of glutamic acid presented in Table 4 do
not seem to be affected by IMP compounds.
Foods 2020, 9, 251
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Table 4. 5’-nucleotides (AMP, IMP, GMP, inosine, and hypoxanthine, g/100 g) and free amino acids
(glutamic and aspartic acid, g/100 g) of beef semitendinosus heated at different temperatures and times.
Temperature
Time
AMP
IMP
GMP
Inosine
Hypoxanthine
Glutamic Acid
Aspartic Acid
Control
0.66 b
46.73 a
5.41 b
1.32 c
0.85 c
0.66
0.57
60 ◦ C
70 ◦ C
6h
12 h
6h
12 h
0.53 b
30.94 b
4.22 b
2.90 b
1.24 a
0.62
0.54
0.61 b
30.21 b
6.98 b
3.95 a
1.13 ab
0.61
0.54
1.25 a
35.84 ab
6.04 b
1.45 c
0.82 c
0.66
0.60
1.44 a
39.71 ab
12.38 a
1.74 c
0.90 bc
0.59
0.55
SEM
P Temp
(T)
P Time
(t)
P
T*t
0.08
2.95
0.89
0.14
0.07
0.03
0.03
<0.001
0.024
0.001
<0.001
<0.001
0.899
0.261
<0.001
0.061
<0.001
<0.001
0.968
0.172
0.768
0.542
0.443
0.059
0.012
0.188
0.125
0.379
a–c Different superscript letters within the same row mean significantly different between treatments (p < 0.05).
SEM: standard error of mean (n = 5). AMP: adenosine-5’-monophosphate; IMP: inosine-5’-monophosphate; GMP:
guanosine-5’-monophosphate.
3.3. Equivalent Umami Concentration (EUC)
Synergistic effects measurement has been carried out to determine the taste synergism between
umami amino acids and umami 5’-nucleotides [16], as shown in Table 5. From EUC values, it can
be clearly seen that the synergistic effect between umami compounds was intensified at 70 ◦ C, with
prolonged cooking contributing the most substantial effect even though it has no significance to control.
Sous-vide cooked at 60 ◦ C was less influenced by umami compounds and led to a lower degree of
EUC values, regardless of cooking time. This implies that the temperature plays a central role in
determining the umami taste. However, the EUC’s actual effect on umami taste properties seems to be
limited because the presence of other compounds might also contribute to the umami taste. EUC also
does not completely represent the umami taste in a complex matrix. Therefore, a further affirmation of
the umami taste between sous-vide treatments by using an electronic tongue was carried out.
Table 5. EUC (mg MSG/g) and umami taste (mV) of beef semitendinosus heated at different temperatures
and times.
Temperature
Time
EUC
*Umami
Control
505.03 a
2.40 b
60 ◦ C
70 ◦ C
6h
12 h
6h
12 h
330.84 b
2.36 b
346.34 b
2.35 b
444.05 ab
2.49 b
532.92 a
2.73 a
SEM
P Temp
(T)
P Time
(t)
P
T*t
32.98
0.04
<0.001
<0.001
0.286
<0.001
0.279
0.007
a–b Different superscript letters within the same row mean significantly different between treatments (p < 0.05). SEM:
standard error of mean (n = 5). * Umami is referred to as the umami taste measured by the electronic tongue system.
EUC: equivalent umami concentration.
3.4. Electronic Tongue
The electronic tongue is inspired by the biological system from the human tongue. The sense
of taste from the human tongue is mediated by taste receptor cells, where a bundle of sensors in the
form of 10,000 taste buds are located [12]. Then, taste perception is processed in the brain. Similarly,
the electronic tongue system uses sensor arrays to identify specific taste, which can translate the
chemical substances of food into an analytical signal. The results are then further analyzed through a
chemometric technique to gain final information of the tested samples. In this study, the reason for
using the electronic tongue rather than being examined by sensory panelists is due to the electronic
tongue sensors being able to be calibrated in order to be reliably consistent [12], so as to measure specific
taste objectively. However, this objective cannot be perceived by the human sensory system in which
panelists (in this case, untrained panelists) sometimes confused unintelligible words (e.g., umami).
Other than safety reasons, the electronic tongue can also provide different access to the sensory
evaluation system.
Foods 2020, 9, 251
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In the present study, the umami taste attribute of different sous-vide cooking temperatures and
times can be seen in Table 5. Umami taste was significantly affected by temperature (p < 0.001), cooking
time (p < 0.001), and by the interaction between temperature and time (p = 0.007). There are no
significant differences in the umami taste among all the tested samples, except sous-vide cooked at
70 ◦ C for 12 h cooking time. Empirically, the temperature at 70 ◦ C, regardless of cooking time, showed
higher umami intensity than at 60 ◦ C. These results are in tandem with the EUC result obtained, as
discussed above. Indirectly, this result gives a clear view that umami taste is better at high temperatures.
Although taste is a key issue for sous-vide cooking, only limited studies have highlighted the umami,
for instance, in pork [9,28] and beef [23]. However, those studies were not directly assessing the sensory
parameter by any means but only based upon chemical compounds (e.g., glutamic acid, IMP, and
GMP) that represent umami taste. Thus, further argumentation of umami by sensory evaluation of
sous-vide at different temperatures and times become difficult due to limited or no related information.
3.5. Principal Component Analysis
In order to better visualize the contribution of umami amino acids and umami 5’-nucleotides
on the umami sensory feature, discriminative principal component analysis (PCA) was carried out
(Figure 2). The PCA variables plot (A) and the observation plot (B) projected in the factor space axes-F1
(56.65%) and axes-F2 (32.26%) were well distinguished. In the variables plot, EUC was located in the
middle of axes-F2 and it showed a good correlation to umami (r = 0.738). Nevertheless, EUC scores
are just an estimation based on the formula umami amino acids and 5’-nucleotides to represent the
synergistic fashion of glutamic acid, aspartic acid, GMP, IMP, and AMP, but it does not show true
umami attribute. The EUC score shows a better connection to 5-nucleotides IMP (r = 0.871), AMP
(r = 0.679), and GMP (r = 0.605) than free amino acid aspartic (r = 0.445) and glutamic acid (r = -0.060).
Conversely, electronic tongue umami represented that only AMP (r = 0.912) and GMP (r = 0.912) were
highly correlated, but weakly correlated to IMP (r = 0.333). This unexpected result was supported by
the observation of Yamaguchi and Ninomiya [11], who found that the taste intensity of IMP itself is
weak, but a strong umami taste is regulated with a glutamate presence. They described this as the
synergistic effect between both compounds to produce the umami sensation. We observed separately
the EUC score between IMP and glutamic acid (excluding aspartic acid, GMP, and AMP), but our
result did not fit with the synergistic effect as suggested, because the umami score for sous-vide
cooking at 70 ◦ C for 12 h (data not shown) was shifted to a lower value than control. The score plot (B)
in Figure 2 presents a clear difference in the relative position of samples according to their cooking
conditions. Thus, sous-vide cooked for 12 h at 70 ◦ C was placed at the lower right quadrant that
attributed strong umami.
In our study, glutamic acid was stable throughout the cooking, as shown in Table 4. It had no
difference to control or even to uncooked meat (0.59 g/100g). When it comes to sous-vide cooking,
a thing that comes into question is the concentration of IMP not being consistent through cooking.
Besides the effect of temperature, as discussed earlier in the text, the effect of cooking time also affects
the inconsistency of IMP content although it was not significant (p = 0.061; Table 4). Our result was
also similar to Ishiwatari, Fukuoka, Hamada-Sato, and Sakai [23] in beef and Sasaki, Motoyama, and
Mitsumoto [28] in pork, who found lower IMP with cooking time. As suggested by many researchers,
IMP is an important compound in amplifying the umami of glutamate [16,19,20,29,30]. However, it is
a very crucial occasion to reach umami by synergism between glutamic acid and IMP in cooked meat
because umami for both parties is dependent on the optimal concentration of IMP. Yamaguchi and
Ninomiya [11] also described a similar finding where subjective taste intensity was induced to the
lower value if the IMP proportion was present at too high or too low concentrations to glutamate.
Therefore, further study is necessary for clarifying the two complexes of IMP and glutamic acid for
their synergistic effect, at which point of temperature and time can optimally trigger the umami.
Foods 2020, 9, 251
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Foods 2020, 9, 251
9 of 11
Figure 2.
Principal component
analysis
of umamiamino
amino acids
andand
5’-nucleotides
on umamion
sensory
Figure 2. Principal
component
analysis
of umami
acids
5’-nucleotides
umami sensory
feature in cooked beef semitendinosus (A: variables plot; B: observation plot). Abbreviation in the A
feature in cooked beef semitendinosus ((A): variables plot; (B): observation plot). Abbreviation in
graph: EUC: equivalent umami concentration; GLU: glutamic acid; ASP: aspartic acid; AMP:
the A graph: adenosine-5’-monophosphate;
EUC: equivalent umami
concentration;
GLU: glutamic
acid; ASP: aspartic acid; AMP:
IMP:
inosine-5’-monophosphate;
GMP: guanosine-5’-monophosphate;
hypox: hypoxanthine. In
the B inosine-5’-monophosphate;
graph, the samples are identified as follows:
and 60-12: sous-vide
adenosine-5’-monophosphate;
IMP:
GMP:60-6
guanosine-5’-monophosphate;
cooked at 60 °C for 6 and 12 h; 70-6 and 70-12: sous-vide cooked at 70 °C for 6 and 12 h; 70-30: control,
hypox: hypoxanthine.
In the B graph, the samples are identified as follows: 60-6 and 60-12: sous-vide
cooked at 70 °C for 30 min.
cooked at 60 ◦ C for 6 and 12 h; 70-6 and 70-12: sous-vide cooked at 70 ◦ C for 6 and 12 h; 70-30: control,
glutamic acid was stable throughout the cooking, as shown in Table 4. It had no
cooked at 70 ◦In
C our
for study,
30 min.
difference to control or even to uncooked meat (0.59 g/100g). When it comes to sous-vide cooking, a
thing that comes into question is the concentration of IMP not being consistent through cooking.
4. Conclusions
Besides the effect of temperature, as discussed earlier in the text, the effect of cooking time also affects
the inconsistency
of IMPhas
content
although
it was not
significant
(p =estimation
0.061; Table 4).
Our result
The umami
taste attribute
been
examined
based
on the
method
ofwas
EUC using an
also similar to Ishiwatari, Fukuoka, Hamada-Sato, and Sakai [23] in beef and Sasaki, Motoyama, and
electronic tongue
system. It was shown that in this study that the temperature effect (60 and 70 ◦ C) is
Mitsumoto [28] in pork, who found lower IMP with cooking time. As suggested by many researchers,
much important
the timecompound
effect (30inmin,
6 h, and
12 h). ofThe
most [16,19,20,29,30].
interesting point
to be
IMPthan
is an important
amplifying
the umami
glutamate
However,
it highlighted
◦ C, with
a very
crucialbe
occasion
to reach
by synergism
between
acidthe
andmost
IMP inapparent
cooked
is the umami istaste
could
induced
by umami
sous-vide
cooking
at 70glutamic
attribute
because umami for both parties is dependent on the optimal concentration of IMP. Yamaguchi
achieved aftermeat
prolonged
cooking, affirmed by both EUC and the electronic tongue and their strong
and Ninomiya [11] also described a similar finding where subjective taste intensity was induced to
correlations. the
Furthermore,
correlation
bywas
PCA
best-described
with 5’-nucleotides
than free
lower value if the
IMP proportion
present
at too high or tooumami
low concentrations
to glutamate.
Therefore, further
study is necessary
for is
clarifying
thealternative
two complexestool
of IMP
and glutamic
acid for
amino acids. Therefore,
an electronic
tongue
the best
to replace
conventional
sensory
synergistic effect, at which point of temperature and time can optimally trigger the umami.
assessment intheir
food
processing applications, especially in determining complex taste attributes such
as umami. 4. Conclusions
The umami taste attribute has been examined based on the estimation method of EUC using an
Author Contributions:
Conceptualization,
Y.-H.H.
I.I.;
methodology,
I.I.; software,
S.-T.J. and
electronic tongue
system. It was shown
thatand
in this
study
that the temperature
effect (60I.I.;
andvalidation,
70 °C) is
Y.-H.H.; formalmuch
analysis,
I.I.
and
Y.-H.H.;
investigation,
I.I.
and
Y.-H.H.;
data
curation,
Y.-H.H.;
writing—original
important than the time effect (30 min, 6 h, and 12 h). The most interesting point to be
draft preparation,
I.I. and is
Y.-H.H.;
writing—review
and editing,
S.-T.J.; visualization,
I.I.with
andthe
Y.-H.H.;
highlighted
the umami
taste could be induced
by sous-vide
cooking at 70 °C,
most supervision,
S.-T.J.; Y.-H.H. and
I.I.
contributed
equally
to
this
research
work.
All
authors
have
read
and
agreed
to the published
apparent attribute achieved after prolonged cooking, affirmed by both EUC and the electronic tongue
version of the manuscript.
and their strong correlations. Furthermore, correlation by PCA best-described umami with 5’free amino
acids.
Therefore,
an electronic
tongueand
is the
best alternative
tool to
Funding: This nucleotides
work was than
supported
by the
Korean
Institute
of Planning
Evaluation
for Technology
in Food,
replace conventional
sensory
assessment
in food
applications,
especially in Development
determining
Agriculture, Forestry
and Fisheries
(IPET)
through
theprocessing
Agri-Bioindustry
Technology
Program,
complex
attributes such
umami.
funded by Ministry
of taste
Agriculture,
Foodasand
Rural Affairs (MAFRA) (Project No. 315017-05-1-SB-140), Republic
of Korea.
Author Contributions: Conceptualization, Y.H.H. and I.I.; methodology, I.I.; software, I.I.; validation, S.T.J. and
Y.H.H.; formal analysis, I.I. and Y.H.H.; investigation, I.I. and Y.H.H.; data curation, Y.H.H.; writing—original
Conflicts of Interest: The authors declare no conflict of interest.
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